Effects of the combined arginase and canavanine treatment on leukemic cells in vitro and in vivo.

It was previously demonstrated in in vitro experiments that canavanine (Cav), a natural toxic arginine analogue of plant origin, is a promising candidate for augmenting the antineoplastic effects of arginine starvation. We demonstrated herein that recombinant human arginase, an arginine degrading enzyme, abrogated growth and significantly increased Cav cytotoxicity toward cultured L1210 murine leukemic cells. Cav co-treatment further reduced cells viability in a time-dependent manner and significantly promoted apoptosis induction. In the pilot study we also evaluated for the first time the potential toxicity of the combined arginine deprivation and Cav treatment in healthy mice. Administration of Cav alone or in combination with pegylated cobalt-containing human arginase (Co-hARG) did not evoke any apparent toxic effects in these animals, with no significant behavioural and survival changes after several weeks of the treatment. The therapeutic effects of the combination of Co-hARG and Cav were provisionally evaluated on the highly aggressive murine L1210 leukemia, which is semi-sensitive to arginine deprivation as a monotreatment. Combination of two drugs did not result in significant prolongation of the survival of leukemia-bearing mice. Thus, we have shown that the proposed combinational treatment is rather non-toxic for the animals. It has to be further evaluated in animal studies with alternative tumor models and/or drug doses and treatment modalities.

N-Farnesyloxy-norcantharimide and N-farnesyl-norcantharimide inhibit the progression of leukemia and increase survival days in a syngeneic mouse leukemia model.

This study investigated the anticancer effects of two newly synthesized norcantharidin analogs, N-farnesyloxy-norcantharimide (NOC15) and N-farnesyl-norcantharimide (NC15), in L1210 cells and in a syngeneic mouse leukemia model (L1210 cell line plus DBA/2 mice). We found that the half-maximal inhibitory concentration (IC50) of NOC15 and NC15 on L1210 cells is 1.56 and 2.62 µmol/l, respectively, and that the IC50 of NOC15 and NC15 on human normal lymphoblast is 207.9 and 2569 µmol/l, respectively. In cell cycle analysis, NOC15 could increase the sub-G1 phase, whereas NC15 could induce G2/M arrest. Annexin-V apoptosis assay indicated that both NOC15 and NC15 could induce cell apoptosis. In the syngeneic mouse leukemia model, both NOC15 and NC15 could increase the survival days of mice and decrease the tumor weight. Moreover, both NOC15 and NC15 could retard the increase in peripheral blood leukocyte count due to L1210 cells. In the subcutaneous (s.c.) group, the treatment with NOC15 could retard the decrease in the weight of the liver and the spleen caused by L1210 cells, whereas the treatment with NC15 could retard the decrease in the weight of the spleen caused by L1210 cells. We conclude that the new compounds NOC15 and NC15 have strong anticancer activity and low toxicity both in vitro and in vivo. NOC15 and NC15 may have the potential to be developed into anticancer agents in the future.

Hexaminolevulinate-mediated photodynamic purging of leukemia cells from BM.

Photodynamic therapy (PDT) with porphyrin precursors has been established for tumor treatment. This study aimed at examining applicability of hexaminolevulinate (HAL) for photodynamic purging of leukemic cells from BM grafts and evaluating the clinical relevance of in vitro models. The PDT dose resulting in no colony formation by leukemic cells in vitro, in pure form or in a mixture with BM cells, was insufficient for complete killing of the leukemic cells ex vivo and for the treatment of the leukemia-bearing animals in vivo. The efficacy of HAL-PDT in cell lines in vitro should be verified in clinically relevant in vivo models.

Combined antitumor effect of cyclophosphamide and bromodeoxyuridine in BDF1 mice bearing L1210 ascites tumors.

We investigated the combined effect of cyclophosphamide (CPA) and 5-bromo-2'-deoxyuridine (BrdUrd) both in mice bearing L1210 ascites tumors and in L1210 leukemic cells in vitro. Administration of BrdUrd (100 mg/kg) for 5 consecutive days before a single dose (80 mg/kg) of CPA significantly extended the survival of mice by 158%, compared with CPA alone. BrdUrd administered at daily doses of 100 or 200 mg/kg for 5 consecutive days did not extended the survival of mice. An in vitro MTT assay revealed that BrdUrd enhanced the cytotoxic effect of 4-hydroxycyclophosphamide, an active form of CPA, in the L1210 cells. These results indicate that BrdUrd enhanced the antitumor effect of CPA both in vivo and in vitro.

Comparative prime-boost vaccinations using Semliki Forest virus, adenovirus, and ALVAC vectors demonstrate differences in the generation of a protective central memory CTL response against the P815 tumor.

Tumor-specific Ags are potential target molecules in the therapeutic treatment of cancer. One way to elicit potent immune responses against these Ags is to use recombinant viruses, which activate both the innate and the adaptive arms of the immune system. In this study, we have compared Semliki Forest virus (SFV), adenovirus, and ALVAC (poxvirus) vectors for their capacity to induce CD8(+) T cell responses against the P1A tumor Ag and to elicit protection against subsequent challenge injection of P1A-expressing P815 tumor cells in DBA/2 mice. Both homologous and heterologous prime-boost regimens were studied. In most cases, both higher CD8(+) T cell responses and better tumor protections were observed in mice immunized with heterologous prime-boost regimens, suggesting that the combination of different viral vectors is beneficial for the induction of an effective immune response. However, homologous immunization with SFV provided potent tumor protection despite a rather moderate primary CD8(+) T cell response as compared with mice immunized with recombinant adenovirus. SFV-immunized mice showed a rapid and more extensive expansion of P1A-specific CD8(+) T cells in the tumor-draining lymph node after tumor challenge and had a higher frequency of CD62L(+) P1A-specific T cells in the blood, spleen, and lymph nodes as compared with adenoimmunized mice. Our results indicate that not only the magnitude but in particular the quality of the CD8(+) T cell response correlates with tumor protection.

Schedule-dependent antitumor activity and toxicity of combinations of 5-fluorouracil and cisplatin/carboplatin against L1210 leukemia-bearing mice.

The combination of cisplatin (CDDP) and 5-fluorouracil (5-FU) has been reported to show marked therapeutic effects on experimental tumors and human malignancies, such as head and neck cancer. In these clinical studies, CDDP was administered on day 1 and followed by a 5-d continuous infusion of 5-FU. However, it was repeatedly shown that the sequence of 5-FU followed by CDDP is more active and less toxic in tumor-bearing animals. Thus, the optimal administration schedule of CDDP and 5-FU against L1210 murine leukemia was examined and compared with that of the combination of 5-FU and carboplatin (JM-8). The combinations of 5-FU (days 1 to 5) and CDDP, given either on day 1 or on day 5, showed a similar level of antitumor activity and toxicity. On the other hand, the combinations of 5-FU (days 1 to 5) and JM-8 given on day 5 showed significantly higher antitumor activity and rather less toxicity, as compared with the combinations on the reverse sequence. Thus, the treatment sequence of platinum compounds followed by a 5-d continuous infusion of 5-FU in many clinical studies appears to be extremely favorable to CDDP than JM-8. In addition, pathological examinations of died mice showed that accumulation of ascites and pleural effusion was inhibited most effectively by JM-8, given alone or in combination with 5-FU. These results strongly suggest that the combination of 5-FU followed by JM-8 will be expected to show more excellent antitumor activity against human malignancies, and may be especially useful in patients who are unable to tolerate CDDP-related toxicity.

In vivo growth of mouse leukemia L1210 cells with metabolic alterations in the subunits of ribonucleotide reductase.

Mouse leukemia L1210 cells selected for resistance to drugs targeted specifically at each of the protein subunits of ribonucleotide reductase were studied for their ability to grow in vivo. The life-span of the mice injected with hydroxyurea-resistant L1210 cells, which have elevated levels of the mRNA and protein for the non-heme iron (NHI, R2) subunit of ribonucleotide reductase, was approximately twice that of the mice injected with equal numbers of the parental wild-type L1210 leukemia cells. The life-span of mice injected with the L1210 cells that had alterations in the effector-binding subunit (EB, R1) was considerably shorter than the mice injected with the parental wild-type L1210 cells. These results provide direct evidence that tumor cells with alterations in the properties of ribonucleotide reductase grow differently in vivo, with defined effects on the host mouse that cause either an increased survival time or a decreased survival time compared to the effects of wild-type L1210 leukemia cells on tumor-bearing mice.

Dietary docosahexaenoic acid levels influence the outcome of arabinosylcytosine chemotherapy in L1210 leukemic mice.

The purpose of this study was to investigate whether dietary supplementation with the n-3 fatty acid docosahexaenoic acid (DHA) in combination with arabinosylcytosine (AraC) chemotherapy could prolong the life expectancy of mice bearing L1210 leukemia. The four control diets included rodent chow, a diet containing 5% of a blended oil mimicking the fatty acid composition of rodent chow, and diets containing 5% or 10% fat with safflower oil as the main oil source. The two DHA-supplemented diets provided 1.5% or 3.5% DHA and 5% or 10% total fat, respectively. After tumor cell inoculation, mice were treated with AraC for 10 days. Mice fed the 5% safflower oil diet (30.1 -/+ 4.1 days), but not those fed the 10% safflower oil diet, survived longer than the chow-fed animals (22.1 -/+ 3.1 days, P = 0.05). The 1.5%-/+ DHA diet (average intake 1.8 g DHA/kg/day) was associated with a longer life span (33.3 -/+ 3.4 days, P < 0.01 vs. chow-fed) and no incidence of death due to drug toxicity. Further increasing DHA intake (4.5 g DHA/kg/day) resulted in shortened survival time (26.5 -/+ 2.0 days), increased circulating tumor cell burden, and lowered red blood cell concentrations. These data suggest that a modest level of dietary DHA or linoleic acid supplementation may improve the antineoplastic efficacy of AraC. However, overconsumption of DHA reverses the beneficial effect of DHA intake on drug sensitivity.

Esters of 2-(1-hydroxyalkyl)-1,4-dihydroxy-9,10-anthraquinones with melphalan as multifunctional anticancer agents.

Eight esters of 2-(1-hydroxyalkyl)-1,4-dihydroxy-9,10-anthraquinone with melphalan were prepared and tested for their antitumor activity (S-180) and cytotoxicity. 2-[1-[4-(p-Bis(2-chloroethyl)-aminophenyl)-butanoyloxy]methyl]-1,4-dihydroxy-9,10-anthraquinone and 2-[1-[4-(p-bis(2-chloroethyl)-aminophenyl)-butanoyloxy]ethyl]-1,4-dihydroxy-9,10-anthraquinone showed remarkable antitumor activity (T/C, 265 and 272%).

Potentiatied antitumor effectiveness of combined chemo-immunotherapy with interleukin-12 and 5-fluorouracil of L1210 leukemia in vivo.

In this study we investigated the efficacy of a combination of IL-12 and 5-FU, a chemotherapeutic exerting several immunomodulatory effects, in murine L1210 leukemia. Mice inoculated with 1 x 10(5) leukemia cells were treated with a single dose of 5-FU (50 mg/kg) and seven daily doses of IL-12 (100 ng/dose), and were observed for survival. Treatment with IL-12 or 5-FU given alone produced moderate anti-leukemic effects. However, combination of both drugs resulted in a significant prolongation of mouse survival time. Importantly, there were 70% of long-term (>60 days) survivors among mice treated with both agents simultaneously. Moreover, we observed 100% of long-term survivors when mice were treated with a minimally increased dose of IL-12 (170 ng) in combination with 5-FU (50 mg/kg). The antileukemic effects were completely abrogated in scid/scid mice and in mice depleted of peritoneal macrophages and significantly decreased after administration of anti-CD3+, anti-CD4+ or anti-CD8+ monoclonal antibodies. Administration of anti-NK1.1 antibodies did not decrease the antileukemic effects indicating that NK cells are not important effectors of this treatment regimen. Collectively, these results indicate that the combination of IL-12 and 5-FU is inducing strong antileukemic responses that are dependent on the presence and activity of macrophages and T lymphocytes and warrant further studies of combined chemo-immunotherapy with IL-12.

Antitumor activity of benzamide riboside and its combination with cisplatin and staurosporine.

Benzamide riboside (BR), a new synthetic nucleoside analogue, has demonstrated a potent cytotoxic activity in murine leukemia in vitro. The purpose of the present investigation was to examine the antitumor activity of BR in mice bearing leukemia L1210. The results revealed that BR possesses a potent antitumor activity in vivo. It increases life-span of mice with leukemia. Synergistic cytotoxicity of BR with select DNA damaging agents, cisplatin (cis-Pt) and staurosporine (STP) was examined in MTT chemosensitivity assay, FACS analyses and apoptotic DNA fragmentation on L1210 cells in culture. A simultaneous treatment of leukemia L1210 cells with the combination of BR and STP resulted in synergistic cytotoxicity that correlated with increased apoptotic activity in those cells. On the other hand, treatment of L1210 cells with combination of BR and cis-Pt resulted in antagonistic cytotoxic effect. Finally, to elucidate the synergistic effect of BR and STP in inducing apoptosis, the attention was directed to the activation of cell death processes through various cell cycle signals. This is the first report describing in vivo antitumor activity of BR and its utilization in combination chemotherapy.

Synergistic effects of Actinobacillus suis cells in combination with 5-fluorouracil on mice bearing murine L1210 leukemia cells.

The mean survival rates of female BDF1 mice transplanted intravenously (i.v.) with murine L1210 leukemia cells were significantly prolonged by intraperitoneal (i.p.) pretreatment (before i.v. transplantation) or by i.p. pre- and post-treatment (before and after the i.v. transplantation) with heat-killed Actinobacillus suis cells ATCC 15557 (AS 15557) alone, as compared with untreated L1210-leukemia-cell-bearing control mice. However, significant prolongation of the mean survival rates was not elicited by the i.p. post-treatment with AS 15557 alone. When 5-fluorouracil (5-FU) was applied i.p. to mice receiving post-treatment with AS 15557 alone, the mean survival rates of the L1210-leukemia-cell-bearing mice were significantly prolonged. The antileukemic action of AS 15557, alone or in combination with 5-FU, against L1210 leukemia was superior to that of a streptococcal preparation (OK-432) and was almost the same as of bacillus Calmette-Guérin with or without 5-FU. The results suggest the possibility that the synergism of AS 15557 in combination with 5-FU may be dependent on the relationship between the indirect immunological function of AS 15557 and the direct cytotoxic action of 5-FU on L1210 leukemia cells.

ICRF-187 rescue in etoposide treatment in vivo. A model targeting high-dose topoisomerase II poisons to CNS tumors.

The catalytic cycle of topoisomerase II is the target of some of the most successful antitumor agents used today, e.g. etoposide (VP-16), in the treatment of testicular cancer and small-cell lung cancer. The cell kill mediated by topoisomerase II poisons can be antagonized by distinct drug types. Thus, we have demonstrated etoposide antagonism with the type-II anthracycline aclarubicin, the antimalarial drug chloroquine, and the cardioprotective agent ICRF-187. In other setups, combinations of agonist and antagonists have led to high-dose regimens for counteracting drug resistance. Thus, the exploitation of folinic acid rescue for methotrexate toxicity and the use of mesna to protect against cyclophosphamide toxicity have enabled the use of high-dose methotrexate and cyclophosphamide protocols. Using a similar approach, we have studied possible ways to apply antagonists to topoisomerase II poisons. NDF1-hybrid female mice were treated with the various drugs and drug combinations. Lethality (LD10 and LD50 values) was computed by use of the maximum-likelihood method, and the antitumor effect of the drugs was compared in mice inoculated i.p. with either L1210 cells or Ehrlich ascites tumor cells. In addition, the compounds were tested on L1210 cells inoculated intracranially. The toxicity of the various drugs was evaluated by weight and leukocyte counts. ICRF-187 rescues healthy mice from lethal doses of topoisomerase II poisons. In mice the ICRF-187 LD10 was 500 mg/kg. Within a wide non-toxic dose range (50-250 mg/kg) of ICRF-187 we found protection against m-AMSA and etoposide lethality. Thus, the LD10 of etoposide increased from 34 mg/kg for the single agent to 122 mg/kg for its combination with ICRF-187, corresponding to a 3.6-fold etoposide dose escalation. In contrast, ICRF-187 did not protect against lethal doses of the non-topoisomerase II-directed drug paclitaxel. We further investigated the anti-tumor effect of equitoxic schedules in mice inoculated i.p. with L1210 or Ehrlich ascites tumor cells. The L1210-bearing mice appeared to obtain a larger increase in life span from the etoposide and ICRF-187 combination as compared with etoposide alone, whereas this was not the case in mice inoculated with Ehrlich ascites tumor cells. As the hydrophilic ICRF-187 is not expected to cross the blood-brain barrier, in contrast to the lipophilic etoposide, we investigated the effect of the drug combination in mice inoculated intracranially with L1210 cells. We obtained a significant increase in life span in mice treated with ICRF-187 + etoposide as compared with mice treated with an equitoxic dose of etoposide alone. Thus, there appear to be potential routes by which one can benefit from this antagonism. ICRF-187 is a powerful nontoxic protector against the lethality of the topoisomerase II-directed drugs etoposide and m-AMSA in vivo. A brain tumor model demonstrates the superiority of high-dose etoposide treatment with ICRF-187 protection as compared with etoposide treatment alone. This implies that tumors in the brain can be reached by cytotoxic drug doses and that normal tissues can be protected due to differences in drug transport across the blood-brain barrier. ICRF-187 is therefore a promising lead compound for the development of schedules using high-dose topoisomerase II poisons in the treatment of brain tumors and metastases.

Correlation of DNA reactivity and cytotoxicity of a new class of anticancer agents: aza-anthracenediones.

Doxorubicin and mitoxantrone are carboxyclic anti-cancer drugs that interact with DNA through intercalation. Our laboratory has synthesized a new series of anti-tumor agents, the aza-anthracenediones, which are structurally related to mitoxantrone but contain a heterocyclic, rather than a carbocyclic, chromophore. Both the in vivo and in vitro anti-tumor activities of these compounds were exquisitely sensitive to the positioning of the nitrogen atom within the heterocyclic backbone. Compounds having a 2-aza were 30- to 100-fold more potent than the 1-aza or the di-aza compounds against L1210 cells in vitro. When tested in vivo, the 2-aza-anthracenediones had marked anti-tumor activity, in some cases curative, whereas the 1-aza-anthracenediones had but minimal antitumor activity. To define the importance of the aza positioning on DNA reactivity, spectral shift and gel mobility assays were used. The spectral shift assay suggested that the 2-aza compounds reacted with DNA solely through intercalation whereas the 1-aza-anthracenediones, and mitoxantrone all reacted with DNA through intercalative and non-intercalative processes. The affinity of DNA binding was five to seven times greater for the 2-aza compounds compared to the 1-aza or the di-aza derivatives. The retardation of supercoiled pBR322 DNA mobility in agarose gel electrophoresis further suggested an intercalative type of DNA interaction. Differences in DNA interaction appear related to but can not completely account for differences in cytotoxicity of the aza anthracenediones.

Augmentation of cancer chemotherapy by preinjection of human macrophage colony-stimulating factor in L1210 leukemic cell-inoculated mice.

Human macrophage colony-stimulating factor (hM-CSF) is a potent stimulator of the effector functions of monocytes/macrophages. We investigated the antitumor effects of this factor in CDF1 male mice inoculated with L1210 cells, a mouse B-cell leukemia line. Mice preinoculated with various numbers of L1210 cells on day 0 were given intravenous injections of vehicle (human serum albumin; HSA) (100 micrograms/kg/day) or hM-CSF (20 micrograms/kg/day) for 3 days from day 1. In mice preinoculated with 10(2) L1210 cells but not with 10(3) or more L1210 cells, a marked increment in survival rate was observed with hM-CSF treatment. We next examined the effect of hM-CSF treatment combined with chemotherapy on the survival of mice that had been preinoculated with 10(5) L1210 cells. In our system, the administration of 4.9 mg/kg adriamycin (ADM) alone slightly prolonged survival of the tumor-bearing mice, but all of the mice died within 20 days. When hM-CSF was injected for 3 days before this ADM treatment, the invasion and proliferation of tumor cells in the liver and spleen were markedly inhibited and 50% of the mice were still alive at day 50. We detected inhibitory activity toward L1210 growth in serum of mice administered with hM-CSF, and the degree of the inhibitory activity was correlated with the level of nitrite (NO2-) in the serum. When L1210 cells were co-cultured with peritoneal macrophages from mice intraperitoneally injected with hM-CSF, the uptake of [3H]thymidine in L1210 cells was inhibited. The inhibition was abolished by the addition of NG-monomethyl-L-arginine, an inhibitor of NO2- synthesis, suggesting that the reactive nitrogen oxide intermediate is involved in hM-CSF-induced inhibition of L1210 growth.

Antitumor activity of cis-malonato[(4R,5R)-4,5-bis(aminomethyl)-2- isopropyl-1,3-dioxolane]platinum(II), a new platinum analogue, as an anticancer agent.

The in vitro and in vivo antitumor activity of a new antitumor platinum complex, cis-malonato[(4R, 5R)-4,5- bis(aminomethyl)-2-isopropyl-1,3-dioxolane]platinum(II) (SKI2053R, NSC D644591), were evaluated and compared with those of cisplatin (CDDP) and carboplatin (CBDCA) using murine tumors. SKI 2053R was highly active in vitro against both L1210 murine leukemia and its CDDP-resistant subline, L1210/DDP; the relative resistances were 20.0-, 14.5-, and 2.7-fold for CDDP, CBDCA, and SKI 2053R, respectively. SKI 2053R showed activity comparable with or superior to either CDDP or CBDCA in mice implanted with L1210. In mice implanted with L1210/DDP, as compared with CBDCA, SKI 2053R showed high values for the percentage of treated survivors relative to controls and for numbers of cured mice, whereas CDDP had virtually no activity. In mice implanted with P388, all three drugs were highly active, but the intensity of activity was shown to be ranked in the following order: SKI 2053R > CDDP > CBDCA. The antitumor activity of SKI 2053R against Lewis lung carcinoma was comparable with that of both CDDP and CBDCA. The antitumor activity of SKI 2053R was further investigated against two human tumor xenografts, KATO III (stomach adenocarcinoma) and WiDr (colon adenocarcinoma), implanted s.c. in nude mice and was compared with that of CDDP. In SKI 2053R-treated groups, the time required for a mean tumor weight of 1,000 mg was 33.1 days in KATO III xenografts and 35.0 days in WiDr xenografts as compared with 30.2 and 27.2 days in CDDP-treated groups, respectively. SKI 2053R achieved growth-inhibition rates comparable with those of CDDP against KATO III (65% versus 59%) and WiDr xenografts (64% versus 54%) on day 35. These results indicate that SKI 2053R is an attractive candidate for further development as a clinically useful anticancer drug.

Enhanced antitumor activity and reduced toxicity of 1,3-bis(2-chloroethyl)-1-nitrosourea administered in lipid microspheres to tumor-bearing mice.

Stable lipid microspheres (LM) and lipid nanospheres (LN) with average diameters of 200 nm and 50 nm, respectively, were used to encapsulate an lipophilic antitumor agent, 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU). LM and LN containing BCNU (lipo BCNU and s-lipo BCNU, respectively) were prepared by homogenizing a soybean oil solution of BCNU with egg yolk lecithin, and their antitumor activity via the intravenous route was tested against L1210 leukemia in mice and compared with that of BCNU dissolved in saline. Both lipo-BCNU and s-lipo BCNU showed significantly enhanced antitumor activity with reduced toxicity, when compared with the corresponding doses of BCNU alone. These results suggest that LM and LN may be suitable carriers for lipophilic antitumor agents and may enhance their efficacy.

Identification of vesicle properties that enhance the antitumour activity of liposomal vincristine against murine L1210 leukemia.

The influence of vesicle lipid composition, size and drug-to-lipid ratio on the antitumour activity of liposomal vincristine was assessed in the murine L1210 ascitic leukemia model. A pH gradient-dependent entrapment procedure was used to encapsulate vincristine and allowed such vesicle properties to be independently varied. Free vincristine delivered i.v. at the maximum tolerated dose (2.0 mg/kg) resulted in a 27.8% increase in the life span (ILS) of mice inoculated i.p. with L1210 cells. Encapsulation of the drug in egg phosphatidylcholine/cholesterol vesicles did not significantly increase the antitumour efficacy of vincristine (ILS, 38.9%). In contrast, administration of vincristine entrapped in vesicles composed of distearoylphosphatidylcholine (DSPC)/cholesterol resulted in ILS values as high as 133%. This enhanced antitumour activity of the DSPC/cholesterol formulations was sensitive to the size of the liposomes; increasing the vesicle size from 100 nm to 1 micron decreased the ILS from 133.3% to 55.6% at a drug dose of 2.0 mg/kg. Decreasing the drug-to-lipid ratio from 0.1:1 to 0.05:1 (w/w) had negligible effects on the activity of liposomal vincristine; however, a further decrease in the drug-to-lipid ratio to 0.01:1 (w/w) decreased the antitumour potency at all drug doses studied. Pharmacology studies indicated that the antitumour activities of free and various liposomal forms of vincristine correlated well with the residence time of the drug in the circulation. These studies indicate that efforts to enhance the therapeutic activity of vincristine through liposome encapsulation must address not only the circulation lifetime of the vesicle systems but also the capacity of the liposomes to retain entrapped drug in vivo.

Effects of 5-fluorouracil and a combination of tegafur and uracil (UFT) on nucleotide metabolism in L1210 ascites tumor.

Changes in the deoxyribonucleotide pools following a single oral administration of 13 mg/kg of 5-fluorouracil (5-FU) or of 64.8 mg/kg of UFT (a mixed compound of tegafur and uracil) were investigated. We compared their pharmacodynamics and effects on nucleotide metabolism in L1210 ascites tumor on day 3 after intraperitoneal tumor inoculation. The intracellular dTTP pool decreased to half the control level 1-6 hr after the administration of 5-FU. The dTTP pools rapidly recovered after 6 hr. In contrast, UFT kept the intracellular dTTP level to 1/3 to 1/2 of the control level for 24 hr. Either drug elevated the intracellular dATP pools, but decreased dCTP pools. UFT influenced the intracellular dATP and dCTP levels longer than 5-FU. Orally administered UFT seemed to exert a longer and more potent inhibitory effect on thymidylate synthetase than equimolar 5-FU. In view of these results, we suggest that UFT could be a more potent chemotherapeutic drug than 5-FU in oral administration.

Correlation of the in vitro cytotoxicity of ethyldeshydroxysparsomycin and cisplatin with the in vivo antitumour activity in murine L1210 leukaemia and two resistant L1210 subclones.

The cultured murine leukaemia L1210 cell populations used in the present study were derived from L1210 cells that had been grown in vivo. Subclones resistant to sparsomycin (L1210/Sm) or cisplatin (L1210/CDDP) were also developed in vivo. The doubling times of the cultured cell populations were identical. Fractions surviving after drug treatment in vitro were determined by colony formation in soft agar. The results, based on the differential sensitivity of the cell populations to ethyldeshydroxysparsomycin (EdSm) and CDDP, indicated that after a short exposure, cultured L1210/CDDP cells were cross-resistant to EdSm. L1210/Sm cells, however, were not cross-resistant to CDDP. The results obtained in cultured cell populations were confirmed in vivo. CD2f1 mice bearing i.p. implants of 1 x 10(5) tumour cells were given EdSm or CDDP and a combination of the two agents. Drugs were given once daily every 4 days for 3 doses starting at 24 h after tumour implantation. Treatment of mice bearing L1210/wt leukaemia with combined EdSm and CDDP caused strongly synergistic antitumour activity. In animals bearing the two resistant subclones, however, combined drug treatment did not improve the antitumour activity. The corresponding median survival of mice receiving combined drug treatment was 60 days in each group containing 6 mice bearing L1210/wt, with 4-6 cures being noted; 19 days in animals harbouring L1210/Sm, with 2 cures being recorded among 6 mice; and 11 days in mice bearing L1210/CDDP, with no cure being obtained. The results of this study indicate that the synergism resulting from combined treatment with CDDP and EdSm is a function of the cellular properties of the target tumour-cell populations and is independent of host factors.